sp_signalsubtract comprises the steps needed to subtract masked regions from a map in order to focus on the remaining features from that map. There are six steps for this process (five within SPHIRE):

1. Average and low-pass filter a map for segmentation :

2. Locate regions to be subtracted :

We recommend using Chimera's segmentation function “Segger” (Tools → Volume Data → Segment Map).

Note the threshold used when segmenting the map. Group the segments in the that you want to exclude; CTRL-click selects a segment, and SHIFT-CTRL-click adds a segment to the selection. (Mouse controls can be customized under “Options.”)

Hint 1: Make the segmented regions transparent (Regions → Make transparent).

Segmented map

With transparency

Hint 2: Hide regions once you have grouped them (Regions → Hide), in case there are segments which are underneath the selected surface segments.

Save the grouped selection to a file (File → Save selections to .mrc file…).

Partially grouped segments

Final grouped segments

The initial map will not have the same dimensions as the original. Pad the map to the target size at the Chimera command line:

vop resample #4 onGrid #2  

…where #4 is the index of the grouped segmented map, and #2 is the original map. Save the padded map to a new file (File → Save map as…).

As described by Alex Belyy

i. Open density map/mask in Chimera. Check pixel size

ii. Load corresponding .pdb models and fit them into the density (Volume → Fit in map).

iii. Select all .pdb model (Tools → General controls → Model Panel, chose pdbs and click select on the right panel)

iv. Color all .pdb models (Actions → color → XXX)

v. Hide the density and select parts of pdb model that you want to highlight in density. Color the selected part with another color (Actions → color → XXX)

vi. Select all .pdb models. Show the EM density.

vii. Tools → Volume data → Color zone

vii. Specify coloring radius, choose the right EM density file and click color. If coloring is satisfactory, click split maps.

ix. In the volume viewer choose the required density. Save it ​

3. Mask a soft mask of the region to be excluded :

4. Compute re-projections of map to be subtracted, and subtract them from the original images :

5. Compute reconstruction of density remaining after signal-subtraction :

6. Center map of the remaining density :

Tapu Shaikh
Alex Belyy

Category 1:: APPLICATIONS